Liposome Nanoparticle Synthesis Objectives

Liposomes are spherical lipid vesicles with a bilayered membrane structure composed of natural or synthetic amphiphilic lipid molecules. Liposomes have been widely used as pharmaceutical carriers in the past decade because of their unique abilities to (a) encapsulate both hydrophilic and hydrophobic therapeutic agents with high efficiency, (b) protect the encapsulated drugs from undesired effects of external conditions, (c) be functionalized with specific ligands that can target specific cells, tissues, and organs of interest, (d) be coated with inert and biocompatible polymers such as polyethylene glycol, in turn prolonging the liposome circulation half-life in vivo, and (e) form desired formulations with needed composition, size, surface charge, and other properties. n this pro!ect, we aim to understand the self-assembly mechanism of lipid molecules to form liposomes and to investigate the preparation of liposomes through an extrusion method. "pecifically, the goals of this pro!ect are# $o prepare liposomes using an extrusion method $o control the size and size distribution of liposomes $o evaluate the quality of the synthesized liposomes $o understand the self-assembly mechanism of the formation of liposomes

References
%hang, L.& 'ranic(, "., )ow to stabilize phospholipid liposome (using nanoparticles). Nano Lett. 2006, *, *+,--. %hang, L.& 'u, .. /.& 0han, 1. 2.& 3ang, 4. %.& Langer, 5. ".& .aro(hzad, 6. 0., 7anoparticles in medicine# therapeutic applications and developments. Clin. Pharmacol. Ther. 2008, -8 (9), :*;-+. $orchilin, <. =., 5ecent advances with liposomes as pharmaceutical carriers. Nat. Rev. Drug Discov. 2005, , (>), ;,9-*?. @arenholz, A., Liposome application# problems and prospects. Curr. Opin. Coll. Inter. Sci. 2001, *, **-::. 2ayer, L. B.& )ope, 2. 1.& 0ullis, =. 5., <esicles of variable sizes produced by a rapid extrusion procedure. Biochim. Biophys. Acta 1 86, -9- (;), ;*;--. Aang, B. 5.& =ornpattananang(ul, B.& 7a(atsu!i, $.& 0han, 2.& 0arson, B.& )uang, 0. 2.& %hang, L., $he antimicrobial activity of liposomal lauric acids against =ropionibacterium acnes. Biomaterials 200 , 8? (8?), *?89-,?.

!reparation
@elow are some (ey steps of the preparation of C;?? nm liposomes using an extrusion method# Bissolve lipids in chloroform at a desired concentration (for example C;,9,;? mgDml, > mL volume)& Evaporate chloroform using nitrogen to purge the solvent (insert the pipette tip inside the chloroform, 8? min for > ml volume) 4dd =@" buffer or B water to rehydrate the lipid film (> mL). <ortex and bath sonicate to accelerate rehydration. (9 min, fully dissolved)& Fse 2ini-Extruder to extrude the large unilamellar vesicles through ;?? nm =0 membranes for at least ;; times & 'et ;?? nm liposomes Fse BL" to obtain size distribution.

"eneral "#i$elines
t is critical to evaporate all organic solvent. 4ny solvent residues will affect the hydration of lipid films when adding =@" buffer, which subsequently decreases the possibility of forming unilamellar lipid vesicles. t is important to do the extrusion process at temperature above the phase transition temperature of the lipid molecules. Lipid molecules usually have a main phase transition temperature, $m. 3hen the environmental temperature is higher than $m, they stay in liquid phase that displays fluidity. n contrast, when the environmental temperature is lower than $m, the lipid molecules stay in gel phase that reflects crystallization of lipid carbon chains. Bo not use the same =0 membrane for more than 8 mL of solution. 6therwise, the membrane may brea( and thus affect the size and size distribution of the resulting liposomes.

Stora%e (for the lab manager)

Lyopholization for storage (G 8 months) HH =repare a second vial and heat it first to (ill bacteriaHH .ilter the liposome suspension using a sterile filter (>?? nm) "tore the filtered liposome solutions in the second vial 4dd ;?I sugar before lyopholization Lyopholize the samples $o use these lyophilized liposomes, after adding buffer, one needs to sonicate them for a couple of minutes to dissolve them well.

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